Several arenaviruses cause hemorrhagic fever (HF) disease in humans and pose a serious threat to public health within their endemic regions. Thus, the New World arenavirus Junin (JUNV) is the etiological agent of Argentine HF (AHF), a severe illness with hemorrhagic and neurological manifestations and a case fatality of 15-30%. In addition, JUNV is very stable, highly infectious by aerosol and causes high morbidity and mortality at low dose, features that make it highly suitable as potential biological weapon. Concerns about arenavirus infections are aggravated by the lack of licensed vaccines in the US and current anti-arenavirus therapies being limited to the use of the nucleoside analogue ribavirin, which is only partially effective and can cause significant toxicity. Accordingly, the development of antiviral drugs against JUNV has been considered as one of the top priorities within the Implementation Plan of the HHS Public Health Emergency Medical Countermeasures Enterprise (PHEMCE). Therefore, medical countermeasures against JUNV infections are eligible for acquisition by BARDA for the Strategic National Stockpile. We have developed a reverse genetic system for JUNV and rescued infectious viruses, both Candid1 and the pathogenic Romero strain, entirely from cloned cDNAs. This advance has provided us with the unique opportunity to generate a genetically well-defined seeds of Candid1 (rJUNV/Candid1cSD) and Romero (rJUNV/Romero) strains of JUNV to be utilized in immunogenicity and efficacy studies. This proposal is designed to provide a comprehensive characterization of the stability, immunogenic and protective properties of rJUNV/Candid1cSD as a first and necessary step for the development of a Candid 1-based vaccine able to meet the safety and efficacy criteria required for future FDA licensing. For this we propose the following specific aims: 1. Determine the genetic and phenotypic stability of rJUNV/Candid1cSD. We propose to generate and characterize a Master Virus Seed (MVS) of rCandid1cSD. We will use the MVS from rCandid1cSD to conduct independent serial passages in different cell substrates, including FDA approved cell lines for vaccine production. Infectious progenies from selected passages from each series will be characterized genetically (genome sequencing) and phenotypically (growth properties and virulence). Genetic and phenotypic information derived from these studies will be used to generate novel rCandid1 strains optimized for genetic and phenotypic stability. 2. To determine the safety, immunogenicity and efficacy of rJUNV/Candid1cSD in guinea pigs. We propose to conduct a comprehensive assessment of the virulence of our genetically well-defined MVS rCandid1cSD in guinea pigs. For this, we will determine clinical parameters associated with immunization of rCandid1cSD including: body temperature and weight, overall activity, assessment of hematological and clinical pathology data covering kidney and liver function, and protection against lethal challenge with JUNV. These studies will also include relevant viral variants identified during studies aimed at assessing the genetic and phenotypic stability of the MVS. 3. Determine the efficacy of rJUNV/Candid1cSD against JUNV in common marmoset (Callithrix jacchus). We propose to evaluate the protective efficacy of rJUNV/Candid1cSD against HF disease caused by the pathogenic rJUNV/Romero strain in the common marmoset (Callithrix jacchus) animal model of JUNV infection. Results from these studies will be correlated with immunogenicity data obtained from studies described in aim 2.